Previous work under this grant has defined two forms of peripheral tolerance that can develop in CDST cells. Naive CDST cells require three signals for full activation;antigen, costimulatory ligand (usually 67-1,2) and either IL-12 or Type IIFN. Antigen recognition in the absence of a 'signal 3'cytokine stimulates proliferation, but the differentiation program leading to effector function and memory does not occur, and the cells are tolerant long-term. Under Aim 1, these 'signal 3 tolerant'cells will be further characterized with respect to their phenotype and functional capacity. Under Aim 2, experiments will be done to determine the mechanistic basis for the non-responsiveness. Preliminary evidence strongly suggests that the non-responsiveness results from a failure of critical gene loci to undergo chromatin remodeling in the absence of a signal from IL- 12/IFN-a. A second form of tolerance can occur when CDST cells become fully activated, but enter a state we have termed activation-induced non-responsiveness (AINR) in which they lose the ability to produce IL-2 to support continued expansion (a form of anergy). Under Aim 3, experiments will be done to determine when the conversion from AINR to responsive memory cells occurs, and define the mechanistic basis for this conversion. It is expected that the results of these studies will contribute to a better understanding of the basic mechanisms responsible for inducing peripheral tolerance in CDST cells, and suggest ways of avoiding or reversing tolerance for therapy of tumors and viral diseases.